An enzyme-linked immunosorbent assay (ELISA) for the determination of mucin levels in bronchoalveolar lavage fluid

Phillips, Jonathan E.; Case, Natalie R.; Celly, Chander; Chapman, Richard W.; Hey, John A.; Minnicozzi, Michael

doi:10.1016/j.vascn.2005.03.001

Cited by 12 publications

(9 citation statements)

References 39 publications

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“…All solutions were maintained at 37°C and continuously equilibrated with 100% O 2 . Enzyme-linked immunosorbent assay was performed as described previously by Phillips et al (2006). Porcine stomach mucin (SigmaAldrich) was used as a standard starting from a concentration of 100 g/ml and then serially diluted to 1:1024.…”

Section: Methodsmentioning

confidence: 99%

Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells

Liu²,

Farley³

2010

J Pharmacol Exp Ther

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Macrolide antibiotics such as erythromycin (EM) and azithromycin (AZM) are beneficial in the treatment of mucus hypersecretion in inflammatory pulmonary diseases. Several indirect and direct mechanisms of action have been proposed. This study investigates the direct effect of macrolides on secretory function of isolated submucosal mucous gland cells (SMGCs). We hypothesize that macrolides inhibit the calcium influx necessary for evoked mucus secretion. To test this, we quantified mucin protein release using enzyme-linked immunosorbent assay, calcium-activated K ϩ (K Ca ), and calcium-activated Cl Ϫ (Cl Ca ) currents. We measured nonselective cation current (NSCC) using whole-cell patch-clamp techniques; intracellular calcium concentration ([Ca 2ϩ ] i ) was measured using fura-2 Ca 2ϩ imaging. We found that both EM and AZM are agonists at muscarinic receptors. EM (10 M) evoked a small but significant increase in mucin release but inhibited the mucin release induced by subsequent acetylcholine (ACh) treatment. Both EM and AZM (10 M) evoked K Ca and Cl Ca whole-cell currents, which were blocked by atropine. EM and AZM also accelerated the decay of inositol trisphosphate-induced K Ca and Cl Ca currents without changing the peak current amplitudes. Likewise, internal application of AZM (10 M) enhanced the decay rate of ACh-induced K Ca and Cl Ca currents. EM (1-10 M) and AZM (0.1-10 M) slowly (over 25-30 min) inhibited thapsigargin (TG)-induced Ca 2ϩ entry when applied during the plateau phase of Ca 2ϩ entry but blunted TG-induced Ca 2ϩ entry by 70% after a 5-min pretreatment before initiating calcium entry. EM blocked TG-induced NSCC. We conclude that macrolide antibiotics are partial agonists at muscarinic receptors but inhibit stimulated mucus release by inhibiting calcium entry in SMGCs.

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Section: Methodsmentioning

confidence: 99%

Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells

Liu²,

Farley³

2010

J Pharmacol Exp Ther

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“…Culture media were collected and determined by ELISA assay using 45M1 antibody on the basis of protocol described elsewhere (24). Purified human tracheal mucin was used as standard as described in our previous study (3).…”

Section: Elisamentioning

confidence: 99%

Vanadium pentoxide (V₂O₅) induced mucin production by airway epithelium

Walters

Zhu

et al. 2011

American Journal of Physiology-Lung Cellular and Molecular Phys

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“…Twenty-four hours after the LPS or vehicle challenge, the rats were euthanized, a tracheal catheter was inserted, and a BAL fluid sample was obtained. The BAL sample was analyzed for total and differential inflammatory cell numbers, for measurement of mucin content using the monoclonal anti-mucin antibody A10G5 (Phillips et al, 2006), and for the measurement of CINC-2 ␣ and CINC-3 by ELISA (IBL-America, Minneapolis, MN). The details for the LPS challenge and BAL cell enumeration have been described previously (Spond et al, 2001).…”

Section: Sch527123 Inhibits Lung Inflammation and Mucus Secretion 487mentioning

confidence: 99%

A Novel, Orally Active CXCR1/2 Receptor Antagonist, Sch527123, Inhibits Neutrophil Recruitment, Mucus Production, and Goblet Cell Hyperplasia in Animal Models of Pulmonary Inflammation

Chapman¹,

Minnicozzi²,

Celly³

et al. 2007

J Pharmacol Exp Ther

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129

104

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Sch527123 [2-hydroxy-N,N-dimethyl-3-[[2-[[1(R)-(5-methyl-2-furanyl)propyl]amino]-3,4-dioxo-1-cyclobuten-1-yl]amino]benzamide] is a potent, selective antagonist of the human CXCR1 and CXCR2 receptors (Gonsiorek et al., 2007). Here we describe its pharmacologic properties at rodent CXCR2 and at the CXCR1 and CXCR2 receptors in the cynomolgus monkey, as well as its in vivo activity in models demonstrating prominent pulmonary neutrophilia, goblet cell hyperplasia, and mucus production. Sch527123 bound with high affinity to the CXCR2 receptors of mouse (K d ϭ 0.20 nM), rat (K d ϭ 0.20 nM), and cynomolgus monkey (K d ϭ 0.08 nM) and was a potent antagonist of CXCR2-mediated chemotaxis (IC 50 ϳ3-6 nM). In contrast, Sch527123 bound to cynomolgus CXCR1 with lesser affinity (K d ϭ 41 nM) and weakly inhibited cynomolgus CXCR1-mediated chemotaxis (IC 50 ϳ1000 nM). Oral treatment with Sch527123 blocked pulmonary neutrophilia (ED 50 ϭ 1.2 mg/kg) and goblet cell hyperplasia (32-38% inhibition at 1-3 mg/kg) in mice following the intranasal lipopolysaccharide (LPS) administration. In rats, Sch527123 suppressed the pulmonary neutrophilia (ED 50 ϭ 1.8 mg/kg) and increase in bronchoalveolar lavage (BAL) mucin content (ED 50 ϭ Ͻ0.1 mg/kg) induced by intratracheal (i.t.) LPS. Sch527123 also suppressed the pulmonary neutrophilia (ED 50 ϭ 1.3 mg/kg), goblet cell hyperplasia (ED 50 ϭ 0.7 mg/kg), and increase in BAL mucin content (ED 50 ϭ Ͻ1 mg/kg) in rats after i.t. administration of vanadium pentoxide. In cynomolgus monkeys, Sch527123 reduced the pulmonary neutrophilia induced by repeat bronchoscopy and lavage (ED 50 ϭ 0.3 mg/kg). Therefore, Sch527123 may offer benefit for the treatment of inflammatory lung disorders in which pulmonary neutrophilia and mucus hypersecretion are important components of the underlying disease pathology.Chronic obstructive pulmonary disease (COPD) is the fourth major cause of death in the United States and is characterized by irreversible airflow limitation due to chronic bronchitis or emphysema (Barnes and Stockley, 2005). The pathological hallmarks of COPD include peripheral airway inflammation dominated by neutrophils, the destruction of the lung parenchyma, submucosal gland hypertrophy, and goblet cell hyperplasia, as well as an increase in proinflammatory cytokines and chemokines (Barnes and Stockley, 2005). Current therapies for COPD are similar to those for asthma and include the use of ␤-adrenoceptor agonists, theophylline, muscarinic antagonists, and corticosteroids (Barnes and Stockley, 2005). However, these treatments do not prevent the progressive decline in lung function and demonstrate clinical activity in only a subpopulation of COPD patients. There is a strong correlation between disease Article, publication date, and citation information can be found at

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An enzyme-linked immunosorbent assay (ELISA) for the determination of mucin levels in bronchoalveolar lavage fluid

Cited by 12 publications

References 39 publications

Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells

Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells

Vanadium pentoxide (V₂O₅) induced mucin production by airway epithelium

A Novel, Orally Active CXCR1/2 Receptor Antagonist, Sch527123, Inhibits Neutrophil Recruitment, Mucus Production, and Goblet Cell Hyperplasia in Animal Models of Pulmonary Inflammation

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An enzyme-linked immunosorbent assay (ELISA) for the determination of mucin levels in bronchoalveolar lavage fluid

Cited by 12 publications

References 39 publications

Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells

Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells

Vanadium pentoxide (V2O5) induced mucin production by airway epithelium

A Novel, Orally Active CXCR1/2 Receptor Antagonist, Sch527123, Inhibits Neutrophil Recruitment, Mucus Production, and Goblet Cell Hyperplasia in Animal Models of Pulmonary Inflammation

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Vanadium pentoxide (V₂O₅) induced mucin production by airway epithelium